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3A) was assembled, using the one-step SLIC method 60 , from five PCR fragments generated from suitable plasmid templates with 1525 bp mutual overlaps, thus introducing the following changes compared with the preceding pSB15 vector: (i) replacement of the constitutive pro S promoter for the engineered PylRS gene by the inducible ara BAD promoter, including the ara C repressor gene, (ii) replacement of the tet p/o for the POI-encoding region, here sfGFPa39, by the lac UV5 p/o , including the lac I repressor gene, and (iii) removal of the tet R repressor cistron downstream of the bla ( amp r ) gene and replacement by the coding region for chloramphenicol acetyltransferase ( cam r ) further harboring an amber stop codon at amino acid position 112